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- PDB-6wo1: Hybrid acetohydroxyacid synthase complex structure with Cryptococ... -

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Basic information

Entry
Database: PDB / ID: 6wo1
TitleHybrid acetohydroxyacid synthase complex structure with Cryptococcus neoformans AHAS catalytic subunit and Saccharomyces cerevisiae AHAS regulatory subunit
Components(Acetohydroxyacid synthase ...Acetolactate synthase) x 2
KeywordsTRANSFERASE / AHAS / pyruvate / FAD / dioxygen
Function / homology
Function and homology information


acetolactate synthase regulator activity / acetolactate synthase complex / acetolactate synthase activity / acetolactate synthase / branched-chain amino acid biosynthetic process / valine biosynthetic process / isoleucine biosynthetic process / thiamine pyrophosphate binding / mitochondrial nucleoid / enzyme regulator activity ...acetolactate synthase regulator activity / acetolactate synthase complex / acetolactate synthase activity / acetolactate synthase / branched-chain amino acid biosynthetic process / valine biosynthetic process / isoleucine biosynthetic process / thiamine pyrophosphate binding / mitochondrial nucleoid / enzyme regulator activity / flavin adenine dinucleotide binding / magnesium ion binding / mitochondrion
Similarity search - Function
Acetolactate synthase, small subunit, C-terminal / Small subunit of acetolactate synthase / Acetolactate synthase, small subunit / AHAS, ACT domain / Acetolactate synthase/Transcription factor NikR, C-terminal / Acetolactate synthase, large subunit, biosynthetic / Acetolactate synthase large subunit, TPP binding domain / ACT domain / TPP-binding enzyme, conserved site / Thiamine pyrophosphate enzymes signature. ...Acetolactate synthase, small subunit, C-terminal / Small subunit of acetolactate synthase / Acetolactate synthase, small subunit / AHAS, ACT domain / Acetolactate synthase/Transcription factor NikR, C-terminal / Acetolactate synthase, large subunit, biosynthetic / Acetolactate synthase large subunit, TPP binding domain / ACT domain / TPP-binding enzyme, conserved site / Thiamine pyrophosphate enzymes signature. / ACT domain profile. / ACT domain / Thiamine pyrophosphate enzyme, central domain / Thiamine pyrophosphate enzyme, central domain / ACT-like domain / Thiamine pyrophosphate enzyme, N-terminal TPP-binding domain / Thiamine pyrophosphate enzyme, N-terminal TPP binding domain / Thiamine pyrophosphate enzyme, C-terminal TPP-binding / Thiamine pyrophosphate enzyme, C-terminal TPP binding domain / Thiamin diphosphate-binding fold / DHS-like NAD/FAD-binding domain superfamily
Similarity search - Domain/homology
2-methylpyrimidin-4-amine / DIPHOSPHATE / FLAVIN-ADENINE DINUCLEOTIDE / VALINE / Acetohydroxy-acid synthase small subunit / Acetolactate synthase small subunit, mitochondrial / Acetolactate synthase
Similarity search - Component
Biological speciesCryptococcus neoformans (fungus)
Saccharomyces cerevisiae (brewer's yeast)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.3 Å
AuthorsGuddat, L.W. / Lonhienne, T.
Funding support Australia, 1items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia)Grant numbers 1087713 & 1147297 Australia
CitationJournal: Nature / Year: 2020
Title: Structures of fungal and plant acetohydroxyacid synthases.
Authors: Thierry Lonhienne / Yu Shang Low / Mario D Garcia / Tristan Croll / Yan Gao / Quan Wang / Lou Brillault / Craig M Williams / James A Fraser / Ross P McGeary / Nicholas P West / Michael J ...Authors: Thierry Lonhienne / Yu Shang Low / Mario D Garcia / Tristan Croll / Yan Gao / Quan Wang / Lou Brillault / Craig M Williams / James A Fraser / Ross P McGeary / Nicholas P West / Michael J Landsberg / Zihe Rao / Gerhard Schenk / Luke W Guddat /
Abstract: Acetohydroxyacid synthase (AHAS), also known as acetolactate synthase, is a flavin adenine dinucleotide-, thiamine diphosphate- and magnesium-dependent enzyme that catalyses the first step in the ...Acetohydroxyacid synthase (AHAS), also known as acetolactate synthase, is a flavin adenine dinucleotide-, thiamine diphosphate- and magnesium-dependent enzyme that catalyses the first step in the biosynthesis of branched-chain amino acids. It is the target for more than 50 commercial herbicides. AHAS requires both catalytic and regulatory subunits for maximal activity and functionality. Here we describe structures of the hexadecameric AHAS complexes of Saccharomyces cerevisiae and dodecameric AHAS complexes of Arabidopsis thaliana. We found that the regulatory subunits of these AHAS complexes form a core to which the catalytic subunit dimers are attached, adopting the shape of a Maltese cross. The structures show how the catalytic and regulatory subunits communicate with each other to provide a pathway for activation and for feedback inhibition by branched-chain amino acids. We also show that the AHAS complex of Mycobacterium tuberculosis adopts a similar structure, thus demonstrating that the overall AHAS architecture is conserved across kingdoms.
History
DepositionApr 23, 2020Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 15, 2020Provider: repository / Type: Initial release
Revision 1.1Jul 22, 2020Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Oct 21, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.3Jan 27, 2021Group: Database references / Category: citation_author / Item: _citation_author.identifier_ORCID
Revision 1.4Oct 18, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Acetohydroxyacid synthase catalytic subunit
B: Acetohydroxyacid synthase regulatory subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)113,8467
Polymers112,6362
Non-polymers1,2105
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)195.679, 195.679, 195.679
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number207
Space group name H-MP432

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Components

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Acetohydroxyacid synthase ... , 2 types, 2 molecules AB

#1: Protein Acetohydroxyacid synthase catalytic subunit


Mass: 78594.672 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cryptococcus neoformans (fungus) / Production host: Escherichia coli (E. coli) / References: UniProt: Q96VZ6, acetolactate synthase
#2: Protein Acetohydroxyacid synthase regulatory subunit


Mass: 34041.734 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Production host: Escherichia coli (E. coli) / References: UniProt: C7GQK9, UniProt: P25605*PLUS

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Non-polymers , 5 types, 5 molecules

#3: Chemical ChemComp-FAD / FLAVIN-ADENINE DINUCLEOTIDE / Flavin adenine dinucleotide


Mass: 785.550 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C27H33N9O15P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: FAD*YM
#4: Chemical ChemComp-8GF / 2-methylpyrimidin-4-amine / 2-methyl-4-aminopyrimidine


Mass: 109.129 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H7N3 / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical ChemComp-DPO / DIPHOSPHATE / Pyrophosphate


Mass: 173.943 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: O7P2 / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#7: Chemical ChemComp-VAL / VALINE / Valine


Type: L-peptide linking / Mass: 117.146 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C5H11NO2 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.77 Å3/Da / Density % sol: 55.62 %
Crystal growTemperature: 291 K / Method: vapor diffusion, hanging drop / Details: Potassium thiocyanate, PEG 3350

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Data collection

DiffractionMean temperature: 100 K / Ambient temp details: Oxford cryostream / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: Australian Synchrotron / Beamline: MX1 / Wavelength: 0.9537 Å
DetectorType: ADSC QUANTUM 210r / Detector: CCD / Date: Mar 28, 2019
RadiationMonochromator: SI(III) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9537 Å / Relative weight: 1
ReflectionResolution: 3.3→48.92 Å / Num. obs: 19890 / % possible obs: 99.8 % / Redundancy: 19.7 % / Biso Wilson estimate: 92.68 Å2 / CC1/2: 0.999 / Rpim(I) all: 0.041 / Net I/σ(I): 13.7
Reflection shellResolution: 3.3→3.56 Å / Num. unique obs: 3985 / Rpim(I) all: 0.286

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Processing

Software
NameVersionClassification
PHENIX1.17.1_3660refinement
XDSdata scaling
PDB_EXTRACT3.25data extraction
XDSdata reduction
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5IMS, 2FGC

5ims

2fgc


Resolution: 3.3→48.92 Å / SU ML: 0.42 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 26.7
RfactorNum. reflection% reflection
Rfree0.2692 1978 9.99 %
Rwork0.2102 --
obs0.2159 19801 99.43 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 253.15 Å2 / Biso mean: 95.6873 Å2 / Biso min: 35.56 Å2
Refinement stepCycle: final / Resolution: 3.3→48.92 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5599 0 79 0 5678
Biso mean--103.31 --
Num. residues----748
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 14

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
3.3-3.380.38141350.31341217135297
3.38-3.470.31271370.237712401377100
3.47-3.570.29791400.232912451385100
3.58-3.690.30231390.23212561395100
3.69-3.820.32391390.240612501389100
3.82-3.970.31291380.240612521390100
3.98-4.160.28631410.208812611402100
4.16-4.370.25681390.186412581397100
4.38-4.650.24061390.18991250138999
4.65-5.010.26071410.17571269141099
5.01-5.510.25181430.199112821425100
5.51-6.310.28371450.232213071452100
6.31-7.940.26181440.20651312145699
7.95-48.920.22631580.19631424158299
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
13.0729-0.2830.27794.3126-0.53884.9847-0.0289-0.0936-0.17860.14250.0542-0.52210.12970.3081-0.03370.5470.01620.15910.5012-0.04920.6305-341.219476.0916353.5143
21.48540.2398-0.05021.21380.11491.14020.00380.48520.0158-0.69420.0503-0.1555-0.1060.0245-0.05440.99860.03950.20440.6876-0.09840.6585-354.1226471.6921326.7126
37.7735-1.2785-0.75932.7879-1.11447.87160.32070.0327-0.1095-0.7743-0.2606-0.32480.3142-0.58690.05460.5861-0.01360.06780.6454-0.06480.5302-367.9908484.6232360.49
43.2912-3.66681.40535.2868-2.44762.11440.47221.7506-0.6373-2.7036-0.23690.95740.9535-1.06650.15281.0107-0.03430.00251.284-0.24680.7692-371.0186480.3286349.9479
52.6153-2.9627-3.28716.31916.18966.1684-0.06880.29830.1007-0.38070.02870.1962-0.1499-0.1710.06770.725-0.01180.15710.66960.16690.728-374.5103496.6776370.8667
65.6411-0.29760.0296.69063.12657.58550.0773-0.06191.01370.27320.07070.2299-0.4960.198-0.03270.65030.00580.04750.69340.11690.6163-379.3606510.5907384.7581
73.1594-1.19340.96958.96364.01159.32180.18610.61180.0102-0.836-0.2132-0.62670.24020.296-0.09580.77790.07640.15760.6790.13240.4937-380.1426496.9442371.8561
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1chain 'A' and (resid 118 through 288 )A118 - 288
2X-RAY DIFFRACTION2chain 'A' and (resid 289 through 714 )A289 - 714
3X-RAY DIFFRACTION3chain 'B' and (resid 74 through 128 )B74 - 128
4X-RAY DIFFRACTION4chain 'B' and (resid 129 through 141 )B129 - 141
5X-RAY DIFFRACTION5chain 'B' and (resid 142 through 180 )B142 - 180
6X-RAY DIFFRACTION6chain 'B' and (resid 181 through 251 )B181 - 251
7X-RAY DIFFRACTION7chain 'B' and (resid 252 through 282 )B252 - 282

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